Regional desynchronization of microglial activity is associated with cognitive decline in Alzheimer’s disease

Background Microglial activation is one hallmark of Alzheimer disease (AD) neuropathology but the impact of the regional interplay of microglia cells in the brain is poorly understood. We hypothesized that microglial activation is regionally synchronized in the healthy brain but experiences regional desynchronization with ongoing neurodegenerative disease. We addressed the existence of a microglia connectome and investigated microglial desynchronization as an AD biomarker. Methods To validate the concept, we performed microglia depletion in mice to test whether interregional correlation coefficients (ICCs) of 18 kDa translocator protein (TSPO)-PET change when microglia are cleared. Next, we evaluated the influence of dysfunctional microglia and AD pathophysiology on TSPO-PET ICCs in the mouse brain, followed by translation to a human AD-continuum dataset. We correlated a personalized microglia desynchronization index with cognitive performance. Finally, we performed single-cell radiotracing (scRadiotracing) in mice to ensure the microglial source of the measured desynchronization. Results Microglia-depleted mice showed a strong ICC reduction in all brain compartments, indicating microglia-specific desynchronization. AD mouse models demonstrated significant reductions of microglial synchronicity, associated with increasing variability of cellular radiotracer uptake in pathologically altered brain regions. Humans within the AD-continuum indicated a stage-depended reduction of microglia synchronicity associated with cognitive decline. scRadiotracing in mice showed that the increased TSPO signal was attributed to microglia. Conclusion Using TSPO-PET imaging of mice with depleted microglia and scRadiotracing in an amyloid model, we provide first evidence that a microglia connectome can be assessed in the mouse brain. Microglia synchronicity is closely associated with cognitive decline in AD and could serve as an independent personalized biomarker for disease progression. Supplementary Information The online version contains supplementary material available at 10.1186/s13024-024-00752-6.

cohorts, the rest are the mice cohorts.p is the p-value of the paired two-sided t-test, d is the corresponding value of Cohen's d (effect size).

Supplementary Fig. S6 Supplementary Fig. S6. Validation analysis using the cognitive dementia rating (CDR) index. (A)
Relationship between DI and CDR sum of boxes (SOB) score for VOIs with significant correlations after FDR correction.Temporal VOIs are indicated by the red box, the rest of the VOIs belong to the parietal lobe.(B) Relationship between the first principal component (PC1) calculated based on the VOIs with significant differences in DI compared to CTRL (Fig. 6) and CDR SOB.p is the correlation p-value (uncorrected), r is the Pearson's r, m is the slope of the fit, b is its intercept.Expression level TSPO expression in Cortex TSPO expression in Hippocampus Expression level Expression level  Supplementary Table S5.Linear fit parameters for DI versus MMSE score for all investigated VOIs.r is the Pearson's r, p is the uncorrected p-value, pFDR is the p-value after FDR correction.Significant correlations (p < 0.05) are shown in italics, correlations that remained significant after FDR correction (pFDR < 0.05) are shown in bold.The regions are sorted according to their p-value.S6.Linear fit parameters for DI versus CDR SOB for all investigated VOIs.r is the Pearson's r, p is the uncorrected p-value, pFDR is the p-value after FDR correction.Significant correlations (p < 0.05) are shown in italics, correlations that remained significant after FDR correction (pFDR < 0.05) are shown in bold.The regions are sorted according to their p-value.

Supplementary Table S6
d b r a i n H i n d b r a i n F o r e b r a i n H i n d b r a i n F o r e b r a i n H i n d b r a i n F o r e b r a i n H i n d b r a i n F o r e b r a i n H i n d b r a i n F o r e b r a i Brain region-and Aβ pathology-specific TSPO expression levels in microglia.(A) Ridge plots showing TSPO expression levels according to genotype and age.In the cortex of APP23 animals, where plaque pathology is pronounced, a subpopulation of microglia expresses high levels of TSPO; this population is enriched in plaque-associated microglia (PAMs; MX04+) both in 17-and 27month-old animals.In the hippocampus, where plaques develop later, a clear increase in TSPO expression only becomes evident in 27-month-old plaque-associated microglial cells.In contrast, no TSPO expression is observed in microglia isolated from the cerebellum, where plaques never develop.(B) Violin plot showing TSPO expression in cortical and hippocampal microglia from 27-month-old APP23 and wildtype animals.* p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001.(C) Density plot showing strongest enrichment for TSPO expression in microglial subpopulations associated with Aβ-pathology.DAM = Disease associated microglia; IRM = Interferon response microglia; HM = Homeostatic microglia.
S9. Microglia desynchronization in acute ischemic stroke model 7 days after surgery.Plots show ICC heatmaps, absolute ICC values and significant connections of the sham mice (n=6) and mice with photothrombotic stroke (n=6).(A) ICC values for all the pairs of the 21 VOIs.(B) Distributions of absolute ICCs, p-values derive from a Wilcoxon signed-rank test.(C) Significant connections (p < 0.005), including cortical (solid line), subcortical (dashed line), and cortical-subcortical (dotted line) connections; the number of the corresponding connections is shown in gray.All significant connections are also projected into 3D brain images, where the color of the connection represents its value; the nodes are individual VOIs, the size of the node reflects the number of its connections; the total number of connections is shown in gray.

Table S4 Cohort Reference cohort p-value DI GLM vs SUVR GLM Cohen's d DI GLM vs SUVR GLM p-value DI BR vs SUVR BR Cohen's d DI BR vs SUVR BR Best region (BR) name
Supplementary TableS2.Overview on training and test cohorts of the human study.CTRL = healthy control; AD = Alzheimer's disease; ANOVA = analysis of variance; SNP = single nucleotide polymorphism; ApoE = Apolipoprotein E; sTREM2 = soluble triggering receptor expressed on myeloid cells type 2; MMSE = mini-mental-state-examination; CDR SOB = cognitive dementia rating sum of boxes Supplementary